4,5-dihydroxypregnanes



United States Patent 4,5-DIHYDROXYPREGNANES Frank B. Colton, Chicago, 111., assignor to G. D. Searle & Co., Chicago, 111., a corporation of Illinois No Drawing. Application August 20, 1954, Serial No. 451,313

8 Claims. (Cl..-260'397.45)

The present application is a Continuation-impart of my copending application $rial No. 440,269,..filech June 29, 1954. It relates to. anew groupnf steroids and, -more specifically, to 4,5,17,2l-tetrahydroxypregnaner3,20=dione, its ll-oxygenated and. normethylderivatives; andesters thereof.

The compounds which constitute myinventioncan be represented by the general structural formula wherein X is a member of the class consisting of-methylene, carbinol and carbonyl radicals, R isa .membenof'the class consisting of hydrogen and lowenhydrocarboncarboxylic acid residues andR is a member ofithe' class -'consisting of hydrogen and methyl; radicals. Amongitthe radicals which R can represent are hydrogen and such lower alkanoyl radicals as formyl, acetyl, propionyl, butyryl, valeryl, hexanoyl, cyclopentanepropionyl, cyclohexaneacetyl, and the. like. Also within the scope of my invention are compounds wherein R represents a benzoyl, toluyl or related aromatic'acyl radical.

The compounds of my invention are conveniently pre-- pared from l7-hydroxycorticosterone, 17-hydroxydesoxycorticosterone, cortisone or theiresters by treatment with osmium tetroxide and hydrogen peroxide in an inert organic solvent. There results the. formation of the corresponding 4,5-dihydroxypregnane derivative. Treatment of a 4,5,l7,21-tetrahydroXy-, 4',5,11B,17;21=pentahydroxy or 4,5,17,2l-tetrahydroXy-ll-oxo derivative of pregnanc- 3,20-dione with an acylatingagent such as an acyl halide or acyl anhydride in pyridine at room temperature causes selective acylation of the hydroxyl groups in the 4-. and 2l-positions, the hydroXyl groups in the 1115-,aridl'l7- positions remaining unafiected. If an lla-hydroxy group is substituted for the 11 fi-hydroxy groupjthe lla-acyloxy derivative can also be formed in preference to ester'ification in the 5-position.

The compounds of myinvention providehi'ghly active pharmacological agents. They are potentlinhibitorsof inflammation and of hypertension. l'l'hewompoundsof my invention are also valuable as intermediates in. organic synthesis. Thus heating ofithe 4,5-dihydroxycompounds with glacial acetic acid and a small amount of p-toluene- Z,7Z7,9 l2 Patented Dec..20, 195.5

.2 sulfonic-.acid,yields.:the-v4 hydroxy-4-pregnenes of the structural formula CHQOR 5 which act as desoxycorticosterone antagonists. The same 4-hydroxypregnene compounds are also obtained under the same conditions from the corresponding 4-acetoXy-5- hydroxy compounds; in this process "I one molecule of acetic acid is eliminated. An alternative procedure for .the synthesis of these 4-hydroxypregnenes from the 4- acyloxy-S-hydroxy and'4,5-dihydroxypregnanes of my invention comprises heating offthese steroids at reflux temperature "for a .few minutes with concentrated hydrochloric acid in methanol.

I have found that-dehydration of the new 4-a cyloxy-5- hydroxysteroids with thionyl chloride in pyridine at 0 C. results in the formation of 5-pregnenes of the structural -formt1la CHzOR .IC=O ---OH On treatment of these S-pregnenes in acetic acid solution with one equivalent of hydrochloric acid at room temperafollowing examples. However, my invention is not to be construed as limited in spirit or in scope by the details set" forth therein. In these examples quantities are indicated as parts by weight. 6 Example] A .mixture of 3.54, parts of 17hydroxydesoxycorti- ;costerone acetate andj280 parts ofv tertiary butanol is .treatedsuccessively -with..a solution ..of '0.38 part of osmium. tetroxide inlG partsof tertiary butanol, 4 parts of a 3Z6l'-N hydrogen'peroxide, solution in tertiary butanol and, after 10 minutes, with 4 additional parts of such a hydrogen peroxide solution. The reaction mixture is collected on a filter, vacuum dried and then applied in a solution ofrethyl acetate in benzene to a chromatography column containing 320 parts of silica gel. The column is washed with 1800 parts of a 10% solution of ethyl acetate in benzene and then with 10,000 parts of 1 15% ethyl acetate in benzene. The column is next eluted with a solution of ethyl acetate in benzene. Evaporation of the eluate and successive recrystallizations from a mixture of ethyl acetate and petroleum ether and from aqueous ethanol yields 4,2l-diacetoxy-S,17-dihydroxypregnane-3,20-dione which melts at about 262264 C. with decomposition. The infrared absorption spectrum shows maxima at about 2.89, 7.29, 8.08, and 9.58 microns. The compound has the structural formula Example 2 To a solution of 1 part of 4,5,17-trihydroxy-21-acetoxypregnane-3,20-dione in 25 parts of methanol there is added slowly one equivalent of 0.1-N aqueous sodium hydroxide. The solution is maintained at room temperature for minutes and then diluted with water and extracted with ethyl acetate. This extract is washed .with concentrated aqueous sodium chloride solution, dried over anhydrous calcium sulfate, filtered and evaporated. The residue contains a mixture of 4,5,17,2l-tetrahydroxypregnane-3,20-dione and 4,17,21-trihydroxy-4pregnene- 3,20-dione which is applied to a silica gel chromatography column. The column is washed with benzene and then with benzene solutions containing increasing proportions of ethyl acetate in benzene. Upon elution with a 15% solution of ethyl acetate in benzene there is first eluted 4,l7,2l-trihydroxy-4-preguane-3,20-dione and then 4,5 ,'l7'

2l-tetrahydroxypregnane-3,ZO-dione. The latter compound does not shown any ultraviolet absorption in the region 240 millimicrons. The infrared absorption spectrum shows a maximum at 2.8 microns and a broad peak at 5.7-5.8 microns. The compound has the structural formula CHzOH =0 CH: L-..

Example 3 A mixture of 37 parts of 17-hydroxydesoxycorticosterone butyrate and 300 parts of tertiary butanol is treated successively with a solution of 3.8 parts of osmium tetroxide in 160 parts of tertiary butanol, 40 parts of a 3.6l-N hydrogen peroxide solution in tertiary butanol and, after 15 minutes, with 40 additional parts of such a hydrogen peroxide solution. The reaction mixture is permitted to stand for 5 days, then concentrated to about one-half of its original volume and extracted with ethyl acetate. This extract is washed successively with water, sodium bisulfite and a saturated sodium chloride solution, dried over anhydrous calcium sulfate, and concentrated in a current of nitrogen to dryness. The residue is dried under vacuum. The 4,5,17-trihydroxy-2l-butyroxypregnane-3,20-dione thus obtained is treated with 500 parts of pyridine in 420 parts of butyric anhydride at room temperature for 24 hours. Then ice is added and the mixture is permitted to stand at room temperature for a day after which the resulting precipitate is collected on a filter, vacuum dried, and dissolved in a 10% solution of ethyl acetate in benzene. This solution is applied to a chromatography column containing silica gel. The column is washed with 5 and 10% solutions of ethyl acetate in benzene and then eluted with an 18% solution of ethyl acetate in benzene. Evaporation of the eluate yields the 4,2l-dibutyroxy-5,17-dihydroxypregnane-3,ZO-dione. The infrared absorption spectrum shows a maximum at 2.8

CHr-CHr-CIEh-CO-O 03 Example 4 A mixture of 10 parts of cortisone acetate and 500 parts of peroxide-free dioxane is treated successively with a solutionof 0.97 parts of osmium tetroxide in 150 parts of dioxane and 40 parts of a 3.6-N hydrogen peroxide solution in anhydrous tertiary butanol. The reaction mixture is permitted to stand at room temperature for 10 days and then treated with water and extracted with ethyl acetate. The extract is washed with saturated aqueous sodium bisulfite solution and then with saturated sodium chloride solution. The solvent is concentrated in vacuo to parts. The residue contains in addition to a small amount of unconverted cortisone acetate,

CHzO-C O-CH:

O OH

This product is mixed at room temperature with 25 parts of pyridine and 25 parts of acetic anhydride. In order to effect complete solution the mixture is warmed for a few minutes on a steam bath. After standing at room temperature for 1 hour, the reaction mixture is treated with ice. The precipitate is collected on a filter and thoroughly washed with water. It is taken up in a 5% solution of ethyl acetate in benzene and applied to a silica gel chromatography column. The column is washed with benzene, a 5% and a solution of ethyl acetate in cenzene and then eluted with a 20% solution of ethyl acetate in benzene. Concentration of this eluate and recrystallization of the residue from ethyl acetate and petroleum ether yields 4,2l-diacetoxy-5,l7-dihydroxypregnane-S',ll,20-trione melting at about 257-259 C. (with decomposition). The infrared absorption spectrum shows maxima at 2.90, 7.28, 8.08, 9.52 and 10.21 microns, and a broad band at 5.7-5.85 microns. The optical rotation of an 0.5% chloroform solution is a =+70. The compound has the structural formula To a stirred mixture of 5 parts of l7-hydroxycorticosterone acetate and 250 parts of peroxide-free dioxane, there are added successively a solution of 0.48 part of osmium tetroxide in 75 parts of dioxane and 40 parts of a 3.6-N hydrogen peroxide solution in anhydrous tertiary butanol. After standing at room temperature for a week, the reaction mixture is treated with water and extracted with ethyl acetate. The extract is washed successively with saturated aqueous sodium bisulfate solution and then with saturated aqueous sodium chloride solution. The solvent is removed under vacuum. The resulting residue contains 4,5,11,17-tetrahydroxy-21- acetoxypregnane-3,20-dione as well as some unconverted starting material. This residue is stirred with parts each of pyridine and acetic anhydride at room temperature for 30 minutes and then treated with ice. A precipitate forms which is collected on a filter, Washed with water, and is then taken up in a solution of 10% ethyl acetate in benzene and thus applied to a silica gel chromatography column. The column is washed with benzene, 5 and 10% solutions of ethyl acetate in benzene, and then eluted with a solution of ethyl acetate in benzene. Concentration of this eluate and recrystallization of the residue from a mixture of ethyl acetate and petroleum ether yields 4,2l-diacetoxy-5,l15,17-trihydroxypregnane-3,20-dione. The ultraviolet absorption shows no maximum in the region of 220 to 250 millimicrons. An infrared maximum is observed at 2.9, broad maxima at 5.7-5.9, and 8.1 microns. The compound has the structural formula CHgQ-C O-GH om-oo-c OH Example 6 A stirred solution of 10.6 parts of 3-methoxy-13- methyl 1,4,6,7,8,9,l1,l2,13,l4,l6,17 dodecahydro 15H cyclopenta[a]phenanthren 17 one (prepared by the process described in my U. S. Patent No. 2,655,518, issued October 13, 1953) in 700 parts of anhydrous ether and 45 parts of dry toluene is cooled to 0 C. and saturated with dry acetylene. While a slow stream of acetylene is passed through the reaction mixture, a solution of 20 parts of potassium t-amylate in 135 parts of anhydrous pentanol is added in the course of 15 minutes With stirring. Passage of acetylene and stirring are continued for an additional 4 /2 hours. After standing at 0 C. for 16 hours, the mixture is washed with aqueous ammonium chloride solution until the aqueous phase is neutral, then with Water and saturated sodium chloride solution. The organic layer is dried over anhydrous sodium sulfate, filtered and concentrated under vacuum to a residue of about 250 parts. 500 parts of petroleum ether are added and, after standing at 0 C. for an hour, the mixture is filtered. The collected precipitate is recrystallized from ether. The resulting 3-methoxy- 13 methyl 17 ethynyl 1,4,6,7,8,9,11,12,13,l4,16,17 dodecahydro- 15H cyclopentaEalphenanthren 17 o1 melts at about 181l82 C. The rotation as determined in a 1% chloroform solution is [a] An additional amount of this product can be obtained from the mother liquors by concentration under vacuum followed by addition of petroleum etherv To a refluxing solution of 47.5 parts of B-methoxy- 13 methyl 17 ethynyl l,4,6,7,8,9,1l,12,l3,l4,l6,17- dodecahydro 15H cyclopenta[a]phenanthren 17 01 in 3200 parts of methanol and 1000 parts of water are added 240 parts of concentrated hydrochloric acid. Refluxing is continued for an additional 5 minutes after which the solution is maintained at room temperature for 15 minutes. Then 13,000 parts of water are added and the mixture is cooled to 0 C. After standing for several hours at that temperature, the mixture is filtered and the precipitate is dried and crystallized from ethyl acetate. The 13 'nethyl l7 ethynyl 17 hydroxy- 1,2,3,6,7,8,9,10,11,12,13,14,l6,17 tetradecahydro 15H- cyclopenta[alphenanthren-3-one thus obtained melts at about 202-204 C. The rotation, as determined in a 1% chloroform solution, is [a] =22.5. The ultraviolet absorption spectrum of a methanolic solution shows a maximum at 241 millimicrons with a molecular extinction coefficient of 17,100.

A solution of 53.7 parts of 13-methyl-l7-ethynyl-l7- hydroxy 1,2,3,6,7,8,9,10,11,12,13,l4,16,17-tetradecahydro-15H-cyclopenta[alphenanthren-3-one in 1500 parts of dioxane and 1000 parts of pyridine is reduced in an atmosphere of hydrogen over 30 parts of a catalyst containing 5% palladium on calcium carbonate. On absorption of one molecule of hydrogen the reduction is stopped and the mixture is filtered. The filtrate is concentrated under vacuum to about 500 parts, diluted with 3000 parts of ether and washed with normal hydrochloric acid until a Congo red test shows an acidic reaction. The solution is washed successively with water, 5% sodium bicarbonate, water and saturated sodium chloride solution. The ether extract is dried over sodium sulfate, concentrated on the steam bath to about 500 parts and diluted with 800 parts of petroleum ether. After storage at C. for 16 hours, the product is collected on a filter, dried and crystallized from a mixture of ethyl acetate and petroleum ether. The 13-methyl-17-vinyl-17- hydroxy 1,2,3,6,7,8,9,10,11,12,13,14,16,17 tetradecahydro-15H-cyclopenta[alphenanthren-3-one thus obtained melts at about 169-171" C. The rotation of an alcoholic solution is [cc] ==+36.

A solution of 47.3 parts of phosphorus tribromide in 645 parts of anhydrous ethanol-free chloroform is added dropwise to a solution of 142.9 parts of 13-methyl-17- vinyl 17 hydroxy-1,2,3,6,7,8,9,10,1l,12,13,14,16,17- tetradecahydro-lSI-I-cyclopenta[a]phenanthren-B-one in 2250 parts of chloroform and 10 parts of pyridine, maintained at -20 C. After standing at room temperature for 16 hours, the mixture is treated with chloroform and then successively with dilute hydrochloric acid, dilute sodium bicarbonate solution and finally with water. After drying over anhydrous sodium sulfate, the chloroform is stripped ofi, leaving as a residue the l7-(fi-bromoethylidene) l3 methyl 1,2,3,6,7,8,9,l0,11,12,13,14, 16,17 tetradecahydro 15H cyc1openta[a]phenanthren- 3-one.

45 parts of 17-(fi-bromoethylidene)-13-methyl-1,2,3, 6,7,8,9,10,11,l2,l3,14,l6,17 tetradecahydro 15H cyclopenta[alphenanthren-3-one are treated with 400 parts of freshly fused potassium acetate and refluxed for 5 hours in 3200 parts of dry acetone. After cooling the precipitate is removed by filtration and the acetone is distilled in vacuum under nitrogen. The residue is extracted by refluxing with boiling petroleum ether and, after stripping of the solvent in vacuo, the residue is chromatographed over 4500 parts of silica gel. Elution with a 3% solution of ethyl acetate in benzene, evaporation of the solvent from the eluate and crystallization of the residue from aqueous acetone and petroleum etheryields 13-methyl-17-vinyl-1,2,3,6,7,8,9,10,11,12,13,14-dodecahydro-l5H-cyclopenta[alphenanthren-3-one, melting at about 100-101" C. The specific rotation of an 0.66% chloroform solution is [a] =+11O.5. The ultraviolet absorption spectrum of a methanolic solution shows. a

maximum at 237 millimicrons with a molecular .extinction coefiicient of 30,200.

Elution of the chromatography column with a solution of ethyl acetate in benzene, evaporation of the solvent from the eluate and recrystallization of the residue from aqueous acetone yields the 17-(B-acetoxyethylidene)-13-methyl-l,2,3,6,7,8,9 ,10,11,12,13,14,16,17-tetradecahydro 15H cyclopentaEalphenanthren 3 one. This compound is obtained in two polymorphic crystalline forms, one melting at 49-50 C., the other melting at about 9697 C. The specific rotation of a 1% chloroform solution is [a] =|62.5. The ultraviolet absorption spectrum of a methanolic solution shows a maximum at 241 millimicrons with a molecular extinction coefllcient of 17,800.

To a solution of 25 parts of 17-(fi-acetoxy-ethylidene)- 13 methyl l,2,3,6,7,8,9,l0,l1,l2,13,14,16,17-tetradecahydro-1SH-cyclopenta[alphenanthren-3-one in 200 parts of tertiary butanol are added 0.27 part of osmium tetroxide in 16 parts of tertiary butanol, followed immediately by 60 parts of a 3.27-N-hydrogen peroxide solution in tertiary butanol. In the course of the following two hours, a solution of 1.25 parts of osmium tetroxide in 80 parts of tertiary butanol is added. After standing at room temperature for 24 hours, the mixture is treated with 1500 parts of water and concentrated in vacuum at room temperature until about 320 parts of distillate have been collected. The residue is extracted with ethyl acetate and the extract is washed with water, dried over sodium sulfate, filtered, and evaporated to dryness. The residue is taken up in 1000 parts of methanol and refluxed for 30 minutes with a solution of 9 parts of sodium sulfite in 200 parts of water. The reaction mixture is concentrated to about one-half of its original volume under nitrogen and extracted with ethyl acetate. This extract is washed with water, dried over sodium sulfate and evaporated. The residue contains a mixture of 13-methyl-l7-glycolyl-17-hydroxy-1,2,3,6, 7,8,9,10,11,12,13,14,16,17 tetradecahydro 15H cyclopenta[a] phenanthren 3 one and 17 (,5 dihydroxyethyl) l7 hydroxy 13 methyl 1,2,3,6,7,8,9, 10,11,12,13,14,16,17 tetradecahydro 15H cyclopentaEalphenanthren-3-one.

In addition the mixture contains athird compound.

which is the 4,5-dihydroxy-13-methyl-17- (p-hydroxyethylidene)perhydro 15H cyclopentaEalphenanthren 3 one of the structural formula The above residue is dissolved in 35 parts of pyridine and 35 parts of acetic anhydride and kept at room temperature for 15 hours. Ice and, 2 hours later, water is added and the mixture is extracted with ethyl acetate. This extract is washed with dilute hydrochloric acid, sodium bicarbonate and water. After drying over sodium sulfate, the extract is evaporated under vacuum and the residue is chromatographed over 550 parts of silica gel. The column is eluted first with 1500 parts of a 10% solution of ethyl acetate in benzene. Elution with 500 parts each of a 10% and a 15% solution of ethyl acetate in benzene yields unreacted starting material. The column is next washed with an additional 500 parts of a 15% solution of ethyl acetate in benzene. Elution with a further 500-part portion of such a 15% solution and evaporation of the solvent yields a residue which, when crystallized from a mixture of ethyl acetate and petroleum ether and then from ether, forms crystals melting at about -187 C. This material gives a positive blue tetrazolium test and does not have a specific absorption in the ultraviolet spectrum between 220 and 330 millimicrons. The infrared spectrum shows maxima at about 2.78, 5.78, 6.9, 7.3, 8.06, 8.79, 9.21, 9.5, 9.75, 10.3, 10.55, 10.8 and 11.3 microns. The compound is the 4-acetoxy-5-hydroxy-13-methyl-17-(fl-acetoxyethylidene)perhydro-1SH-cyclopenta[alphenanthren-3-one of the structural formula 1 CHa CHgO-O C CH3 of CHsCX H Example 7 To a solution of 10 parts of 4acetoxy-5-hydroxy-13- methyl 17 (B acetoxyethylidene)perhydro 15H cyclopenta[alphenanthren-S-one in 75 parts of tertiary butanol are added 0.9 part of osmium tetroxide in 5.3 parts of tertiary butanol followed immediately by 20 parts of a 3.27-N hydrogen peroxide solution in tertiary butanol. A solution of 0.41 part of osmium tetroxide in 26 parts of tertiary butanol is added during the entire reaction time. The reaction mixture is exposed to an efficient source of ultraviolet light. After 24 hours of standing at room temperature 500 parts of water are added and the reaction mixture is concentrated in vacuum until about 100 parts of the distillate have been collected. The residue is extracted with ethyl acetate and the extract is washed with water, dried over anhydrous calcium sulfate, filtered and evaporated to dryness. The residue is taken up in 400 parts of methanol and refluxed for 30 minutes with a solution of 3 parts of sodium sulfite in 80 parts of water. Concentration of the reaction mixture to about one-half of its original volume under nitrogen is followed by extraction with ethyl acetate. This extract is washed with water, dried over anhydrous calcium sulfate and evaporated. The residue contains a mixture of 4-acetoxy-5,17-dihydroxy- 13 methyl 17 glycolylperhydro 15H cyclopenta[a]- phenanthren-3-one of the structural formula CH:|OH

onto o- OH I claim: 1. A compound of the structural formula (IIHQOR a (3:0 Lion X R0 OH wherein X is a member of the class consisting of methylene, carbinol and carbonyl radicals, R is a member (lower alkyD-C o-o 0H 3. 4,21-diacetoxy-5,17-dihydroxypregnane-3,20-dione. 4. 4,5,17,21-tetrahydroxypregnane-3,20-dione.

5. A compound of the structural formula (lower alkyD-CO-O 0H 6. 4,21 diacetoxy 5,17 dihydroxypregnane 3,11, 20-trione.

7. A compound of the structural formula omo-o O-(lower alkyl) =0 L-on (lower a1kyl)-C O- 8. 4,21 diacetoxy 5,11,52,17 trihydroxypregnane 3, ZO-dione.

References Cited in the file of this patent UNITED STATES PATENTS 2,341,081 Butendandt Feb. 8, 1944 FOREIGN PATENTS 497,394 Great Britain Dec. 15, 1938 

1. A COMPOUND OF THE STRUCTURAL FORMULA 